Polymorphic hydroxylation of perhexiline in vitro

Abstract

Aims: The aims of this study were to examine the in vitro enzyme kinetics and CYP isoform selectivity of perhexiline monohydroxylation using human liver microsomes. Methods Conversion of rac-perhexiline to monohydroxyperhexiline by human liver microsomes was assessed using a high-performance liquid chromatography assay with precolumn derivatization to measure the formation rate of the product. Isoform selective inhibitors were used to define the CYP isoform profile of perhexiline monohydroxylation. Results: The rate of perhexiline monohydroxylation with microsomes from 20 livers varied 50-fold. The activity in 18 phenotypic perhexiline extensive metabolizer (PEM) livers varied about five-fold. The apparent Km was 3.3 ± 1.5 μM, the Vmax was 9.1 ± 3.1 pmol min-1 mg-1 microsomal protein and the in vitro intrinsic clearance (Vmax/Km) was 2.9 ± 0.5 μl min-1 mg-1 microsomal protein in the extensive metabolizer livers. The corresponding values in the poor metabolizer livers were: apparent Km 124 ± 141 μM; Vmax 1.4 ± 0.6 pmol min-1 mg-1 microsomal protein; and intrinsic clearance 0.026 μl min-1 mg-1 microsomal protein. Quinidine almost completely inhibited perhexiline monohydroxylation activity, but inhibitors selective for other CYP isoforms had little effect. Conclusions: Perhexiline monohydroxylation is almost exclusively catalysed by CYP2D6 with activities being about 100-fold lower in CYP2D6 poor metabolizers than in extensive metabolizers. The in vitro data predict the in vivo saturable metabolism and pharmacogenetics of perhexiline.